Our long-term objectives are to analyze the organization of the hierarchical controls and the cellular mechanisms involved in a model neuroendocrine system, the bag cell neurons of the opisthobranch mollusk, Aplysia. These 800 neurons (organized into two discrete clusters in the abdominal ganglion) control ovulation and egg-laying behavior by release of a peptide hormone, ELH, during a long lasting (30 min) electrical afterdischarge. The distribution of bag cell processes has been specifically mapped, by a rabbit antibody to ELH, using the peroxidase-antiperoxidase technique, and processes are found inboth neurohaemal areas and neuropile within the ganglion. We have determined the primary structure of ELH which has 36 amino-acid residues. 2.5 nanomoles of ELH is sufficient, by itself, to induce egg laying and correlated behavior in Aplysia. We have also determined that two peptides (A and B) from the atrial gland of the reproductive tract initiate discharge in the bag cells and cause egg-laying through the bag cell system. We have determined the primary structure of these two neuroactive peptides. They each have 34 amino-acid residues, differ from each other in only 4 residues and bear no resemblance, in primary structure, to ELH. There must be receptors for these two atrial gland peptides in the head ganglia for localized application there will trigger bag cell discharge. Recently, a third atrial gland peptide has been isoalted, by another lab, which induces egg-laying, and is a hybrid betwenELH and peptides A and B. However, we have only limited information on crucial aspcts of the overall system and therefore will continue to investigate: 1) The neural nd humoral pathways and the associated chemical factors activating and inhibiting the bag cell system. 2) The time course of ELH release during afterdischarge and its modulation. 3) The use of monoclonal and polyclonal antibodies as pharmacological probes and to localize function-specific peptide and protein production in the bag cells, atrial gland and related systems in the head ganglia. 4) The ionic basis and involvement of protein phosphorylation in electrical afterdischarge in the intact bag cell cluster. Our methodology is broad-based. We use biochemical separation methods (gel filtration, ion exhcnage, gel electrophoresis, HPLC) and other analytical techniques. In collaboration with Drs. Hunkapiller and Hood we obtain amino-acid composition and peptide primary structure. We generate antibodies and can localize their binding with light and electron-microscope techniques. Our electrophysiological techniques include both extracellular and intracellular recording methods.